Bispecific antibodies are multifunctional antibodies that comprise antigen-binding sites that can bind two distinct antigenic determinants and have emerged as one of the major therapeutic biologics for treating many diseases, including cancer. While a variety of bispecific antibodies with dual antigen-binding properties have been developed recently, the specificity and affinity of immunoglobulin light chain or heavy chain variable domains in the conventional bispecific antibodies had to be sacrificed to some extent because, in the conventional bispecific antibodies, either only a heavy chain or a light chain variable domain contributes to binding to each antigenic determinant, whereas, in regular antibodies, both light and heavy chain variable regions can contribute to binding to the same antigenic determinant. In addition, in achieving a desirable level of efficacy, therapeutic antibodies, e.g., bispecific therapeutic antibodies, often require high or multiple doses of antibodies due to their limited recyclability in vivo.
Most antigen-binding proteins that target two antigens or epitopes developed so far comprise two antigen-binding arms: (i) a first antigen-binding arm comprising an immunoglobulin heavy-light chain variable domain pair that contributes to binding to a first antigen or epitope; and (ii) a second antigen-binding arm comprising a second heavy-light chain variable domain pair that contributes to binding to a second antigen or epitope. These antigen-binding proteins, though bispecific in the context of the whole antigen-binding protein, are not necessarily bispecific within each antigen-binding arm, limiting the use of the antigen-binding proteins in multi-specific formats, e.g., tri-specific antigen-binding proteins. As disclosed herein, a non-human animal that expresses a universal heavy chain variable domain may be employed as a general tool for making antigen-binding proteins for use in many different formats of antigen-binding proteins.